Assay for detecting Chlamydia trachomatis, Neisseria gonorrhoeae, Trichomonas vaginalis, and Mycoplasma genitalium

Patent metadata

Field	Value
Publication number	US-11015228-B2
Application number	US-201816237077-A
Country	US
Kind code	B2
Filing date	Dec 31, 2018
Priority date	Jan 9, 2018
Publication date	May 25, 2021
Grant date	May 25, 2021

Abstract

The invention is directed to methods, kits, and compositions, for amplifying and detecting Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), Trichomonas vaginalis (TV), and Mycoplasma genitalium (MG) in a sample, which comprises a variety of combinations of forward oligonucleotide primers, reverse oligonucleotide primers, and oligonucleotide probes.

First claim

The invention claimed is:

1. A set of oligonucleotide sequences for amplifying and detecting Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), Trichomonas vaginalis (TV), and Mycoplasma genitalium (MG) nucleic acid sequences in a sample, which comprises:
(a) a primer and probe set that amplifies and detects at least a portion of the Chlamydia trachomatis 23S rRNA comprising a forward primer oligonucleotide sequence comprising SEQ ID NO: 1, a reverse primer oligonucleotide sequence comprising SEQ ID NO: 2, and a first probe oligonucleotide sequence comprising SEQ ID NO: 3;
(b) a primer and probe set that amplifies and detects at least a portion of the Neisseria gonorrhoeae opa gene comprising a forward primer oligonucleotide sequence comprising SEQ ID NO: 4, a reverse primer oligonucleotide sequence comprising SEQ ID NO: 5, and a second probe oligonucleotide sequence comprising SEQ ID NO: 6;
(c) a primer and probe set that amplifies and detects at least a portion of the Trichomonas vaginalis 18S rRNA comprising a forward primer oligonucleotide sequence comprising SEQ ID NO: 7, a reverse primer oligonucleotide sequence comprising SEQ ID NO: 8, and a third probe oligonucleotide sequence comprising SEQ ID NO: 9; and
(d) a primer and probe set that amplifies and detects at least a portion of the Mycoplasma genitalium 23S rRNA comprising a forward primer oligonucleotide sequence comprising SEQ ID NO: 10, a reverse primer oligonucleotide sequence comprising SEQ ID NO: 11, and a fourth probe oligonucleotide sequence comprising SEQ ID NO: 12, wherein each of the probe oligonucleotide sequences comprises a detectable label.

2. The set of claim 1 , which further comprises an internal control primer and probe set.

3. The set of claim 2 , wherein the internal control primer and probe set comprises:
(e) an internal control forward primer oligonucleotide sequence comprising SEQ ID NO: 13,
(f) an internal control reverse primer oligonucleotide sequence comprising SEQ ID NO: 14, and
(g) an internal control probe oligonucleotide sequence comprising SEQ ID NO: 15 and a detectable label.

4. The set of claim 1 , which further comprises a cellular control primer and probe set.

5. The set of claim 4 , wherein the cellular control primer and probe set amplifies and detects the human β-globin gene.

6. The set of claim 4 , wherein the cellular control primer and probe set comprises:
(h) a cellular control forward primer oligonucleotide sequence comprising SEQ ID NO: 16,
(i) a cellular control reverse primer oligonucleotide sequence comprising SEQ ID NO: 17, and
(j) a cellular control probe oligonucleotide sequence comprising SEQ ID NO: 18 and a detectable label.

7. The set of claim 1 , wherein the detectable label is a fluorophore.

8. The set of claim 1 , wherein each of the probe oligonucleotides further comprises a quencher moiety.

9. A method for detecting Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), Trichomonas vaginalis (TV), and Mycoplasma genitalium (MG) in a sample suspected of containing Chlamydia trachomatis, Neisseria gonorrhoeae, Trichomonas vaginalis , and Mycoplasma genitalium , which method comprises:
(a) contacting a sample obtained from a human with the set of oligonucleotide sequences of claim 1 and reagents for amplification and detection of nucleic acid sequences, and
(b) detecting hybridization of the first, second, third, and/or fourth oligonucleotide probes to an amplified portion of the CT 23S rRNA, the NG opa gene, the TV 18S rRNA, and/or the MG 23S rRNA by assessing a signal from each of the detectable labels, whereby
(i) the presence of a signal from any of the detectable labels indicates the presence of CT, NG, TV, and/or MG in the sample, and
(ii) the absence of a signal from any of the detectable labels indicates the absence of CT, NG, TV, and/or MG in the sample.

10. The method of claim 9 , wherein the sample comprises endocervical tissue or fluid, vaginal tissue or fluid, urine, or cervical tissue or fluid.

11. A kit for detecting Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), Trichomonas vaginalis (TV), and Mycoplasma genitalium (MG) in a sample comprising
(a) a primer and probe set that amplifies and detects at least a portion of the Chlamydia trachomatis 23S rRNA comprising a forward primer oligonucleotide sequence comprising SEQ ID NO: 1, a reverse primer oligonucleotide sequence comprising SEQ ID NO: 2, and a first probe oligonucleotide sequence comprising SEQ ID NO: 3;
(b) a primer and probe set that amplifies and detects at least a portion of the Neisseria gonorrhoeae opa gene comprising a forward primer oligonucleotide sequence comprising SEQ ID NO: 4, a reverse primer oligonucleotide sequence comprising SEQ ID NO: 5, and a second probe oligonucleotide sequence comprising SEQ ID NO: 6;
(c) a primer and probe set that amplifies and detects at least a portion of the Trichomonas vaginalis 18S rRNA comprising a forward primer oligonucleotide sequence comprising SEQ ID NO: 7, a reverse primer oligonucleotide sequence comprising SEQ ID NO: 8, and a third probe oligonucleotide sequence comprising SEQ ID NO: 9;
(d) a primer and probe set that amplifies and detects at least a portion of the Mycoplasma genitalium 23S rRNA comprising a forward primer oligonucleotide sequence comprising SEQ ID NO: 10, a reverse primer oligonucleotide sequence comprising SEQ ID NO: 11, and a fourth probe oligonucleotide sequence comprising SEQ ID NO: 12;
(e) reagents for amplifying and detecting nucleic acid sequences; and
(f) instructions for use,
wherein each of the probe oligonucleotide sequences comprises a detectable label.

12. The kit of claim 11 , which further comprises an internal control primer and probe set.

13. The kit of claim 12 , wherein the internal control primer and probe set comprises:
(e) an internal control forward primer oligonucleotide sequence comprising SEQ ID NO: 13,
(f) an internal control reverse primer oligonucleotide sequence comprising SEQ ID NO: 14, and
(g) an internal control probe oligonucleotide sequence comprising SEQ ID NO: 15 and a detectable label.

14. The kit of claim 11 , which further comprises a cellular control primer and probe set.

15. The kit of claim 14 , wherein the cellular control primer and probe set amplifies and detects the human β-globin gene.

16. The kit of claim 14 , wherein the cellular control primer and probe set comprises:
(h) a cellular control forward primer oligonucleotide sequence comprising SEQ ID NO: 16,
(i) a cellular control reverse primer oligonucleotide sequence comprising SEQ ID NO: 17, and
(j) a cellular control probe oligonucleotide sequence comprising SEQ ID NO: 18 and a detectable label.

17. The kit of claim 11 , wherein the primers, probes, and reagents are lyophilized.

18. A composition for amplifying and detecting Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), Trichomonas vaginalis (TV), and Mycoplasma genitalium (MG) in a sample, which comprises:
(a) a primer and probe set that amplifies and detects at least a portion of the Chlamydia trachomatis 23S rRNA comprising a forward primer oligonucleotide sequence comprising SEQ ID NO: 1, a reverse primer oligonucleotide sequence comprising SEQ ID NO: 2, and a first probe oligonucleotide sequence comprising SEQ ID NO: 3;
(b) a primer and probe set that amplifies and detects at least a portion of the Neisseria gonorrhoeae opa gene comprising a forward primer oligonucleotide sequence comprising SEQ ID NO: 4, a reverse primer oligon

Assignees

• Abbott Molecular Inc

Inventors

• Nelson Kevin

Classifications

• C12Q2600/16

  Primer sets for multiplex assays · CPC title

• C12Q2600/166

  Oligonucleotides used as internal standards, controls or normalisation probes · CPC title

• C12R2001/90

  Protozoa {; Processes using protozoa} · CPC title

• C12R2001/35

  Mycoplasma · CPC title

• C12R2001/01

  Bacteria or Actinomycetales {; using bacteria or Actinomycetales} · CPC title